<2022-07-05 Tue> David biweekly meeting

project updates

Project 4: Reconstitution of endocytic actin network

recent progress

  • Danielle finished sectioning and staining first region from each condition in these slides
  • in the one full day I've had access to the TEM so far, imaged dozens of beads in a single section for the -Las17 -LatA condition

    2022-07-04_18-05-52_Screenshot from 2022-07-04 18-03-22.png
    • typical bead surface:

      2022-07-04_18-51-03_Screenshot from 2022-07-04 18-50-44.png
  • finished analyzing 3D time lapses for budding events in complementary conditions to EM sample conditions

    condition mean budding events per bead standard deviation
    - Las17 - LatA 0.047619047 0.082478608
    + Las17 - LatA 0.24777165 0.29060322
    + Las17 + LatA 0.048082216 0.063811303
    + Las17 - LatA - extract 0 0
    • this is a summary of individual experiments below:

      sample condition total beads budding events events per bead
      4.18.1M1-01 - Las17 - LatA 12 0 0
      4.18.1M2-01 + Las17 - LatA 11 0 0
      4.18.1M2-02 + Las17 - LatA 16 3 0.1875
      4.18.1M3-01 + Las17 + LatA 19 1 0.052631579
      4.18.1M3-02 + Las17 + LatA 30 0 0
      4.18.2M1-01 - Las17 - LatA 23 0 0
      4.18.2M2-01 + Las17 - LatA 31 2 0.064516129
      4.18.2M2-02 + Las17 - LatA 19 14 0.73684211
      4.18.2M3-02 + Las17 + LatA 29 0 0
      4.18.2M3-04 + Las17 + LatA 23 0 0
      4.18.2M3.2-01 + Las17 + LatA 17 3 0.17647059
      4.18.2M3.2-03 + Las17 + LatA 14 0 0
      4.18.2M4-01 + Las17 - LatA - extract 16 0 0
      4.18.3M1-01 - Las17 - LatA 14 2 0.14285714
      4.18.3M2-01 and 02 + Las17 - LatA 20 5 0.25
      4.18.3M3-01 + Las17 + LatA 20 2 0.1
      4.18.3M3-02 + Las17 + LatA 36 2 0.055555556

next steps

  • finish EM imaging and analysis

Nikhil's protein purification project

  1. express and purify His-tagged GFP in E. coli
    • confirmed expression
    • currently scaling up for purification
  2. insert GFP into His6-eps15 plasmid from Stachowiak lab
  3. express and purify His6-eps15-GFP from E. coli
  4. purify His6-eps15
  5. purify Fcho1
  6. reproduce phase separation experiments

Date: \today

Author: Max Ferrin

Email: ferrinm@berkeley.edu

Created: 2022-07-04 Mon 18:51

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